NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss
ABSTRACT This study investigated how NPC1L1, a cholesterol transporter, regulates osteogenic differentiation through cholesterol metabolism independently of its transport function. We also explored the role of NPC1L1 in osteoporosis (OP), focusing on the downstream C/EBPα/Cyp27a1/27‐hydroxycholester...
Saved in:
| Main Authors: | , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2025-06-01
|
| Series: | FASEB BioAdvances |
| Subjects: | |
| Online Access: | https://doi.org/10.1096/fba.2025-00044 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1850215496347025408 |
|---|---|
| author | Bohao Li Wuling Zhou Yueming Yu Boyu Chen Zhicheng Lv Jiarui Zhang Tieqi Zhang Shiwei Sun Lei Zhou Minghai Wang |
| author_facet | Bohao Li Wuling Zhou Yueming Yu Boyu Chen Zhicheng Lv Jiarui Zhang Tieqi Zhang Shiwei Sun Lei Zhou Minghai Wang |
| author_sort | Bohao Li |
| collection | DOAJ |
| description | ABSTRACT This study investigated how NPC1L1, a cholesterol transporter, regulates osteogenic differentiation through cholesterol metabolism independently of its transport function. We also explored the role of NPC1L1 in osteoporosis (OP), focusing on the downstream C/EBPα/Cyp27a1/27‐hydroxycholesterol (27‐OHC) axis. High‐throughput RNA sequencing and bioinformatics analysis identified NPC1L1 as a key regulator of osteogenesis. Osteogenic differentiation assays, Alizarin Red S and ALP staining, western blot analysis, and qRT‐PCR were performed using osteoblast cell lines (C3H10 and C2C12). In addition, an ovariectomy (OVX)‐induced mouse model of OP was established to validate the in vivo effects. ELISAs, chromatin immunoprecipitation (ChIP–qPCR), and rescue experiments were conducted to verify the functional interactions among NPC1L1, Cyp27a1, 27‐OHC production, and the transcription factor C/EBPα. NPC1L1 expression was downregulated during osteogenesis, and its knockdown significantly enhanced osteogenic differentiation, proliferation, and migration. At the molecular level, NPC1L1 promoted cholesterol metabolism independently of its transport function, resulting in elevated 27‐OHC levels through increased expression of Cyp27a1. Elevated 27‐OHC suppressed osteogenesis through the induction of oxidative stress and the downregulation of osteogenic biomarkers (ALP, OPN, OSX, and OCN). In OVX mice, NPC1L1 knockdown significantly reversed osteoporosis‐related bone loss, as evidenced by improved trabecular parameters (BV/TV%, Tb.Th, Tb.N). Furthermore, we identified C/EBPα as a transcriptional activator of Cyp27a1, which mediates the regulatory effects of NPC1L1 on 27‐OHC production. NPC1L1 inhibits osteogenesis and contributes to OP by promoting the Cyp27a1‐dependent synthesis of 27‐OHC through the transcription factor C/EBPα. Targeted modulation of the NPC1L1‐C/EBPα‐Cyp27a1‐27‐OHC axis could provide novel therapeutic strategies for OP. |
| format | Article |
| id | doaj-art-548ec740e42f4fae8466aa6b44d245d0 |
| institution | OA Journals |
| issn | 2573-9832 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Wiley |
| record_format | Article |
| series | FASEB BioAdvances |
| spelling | doaj-art-548ec740e42f4fae8466aa6b44d245d02025-08-20T02:08:36ZengWileyFASEB BioAdvances2573-98322025-06-0176n/an/a10.1096/fba.2025-00044NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone LossBohao Li0Wuling Zhou1Yueming Yu2Boyu Chen3Zhicheng Lv4Jiarui Zhang5Tieqi Zhang6Shiwei Sun7Lei Zhou8Minghai Wang9Department of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaDepartment of Orthopedics, Shanghai Fifth People's Hospital Fudan University Shanghai ChinaABSTRACT This study investigated how NPC1L1, a cholesterol transporter, regulates osteogenic differentiation through cholesterol metabolism independently of its transport function. We also explored the role of NPC1L1 in osteoporosis (OP), focusing on the downstream C/EBPα/Cyp27a1/27‐hydroxycholesterol (27‐OHC) axis. High‐throughput RNA sequencing and bioinformatics analysis identified NPC1L1 as a key regulator of osteogenesis. Osteogenic differentiation assays, Alizarin Red S and ALP staining, western blot analysis, and qRT‐PCR were performed using osteoblast cell lines (C3H10 and C2C12). In addition, an ovariectomy (OVX)‐induced mouse model of OP was established to validate the in vivo effects. ELISAs, chromatin immunoprecipitation (ChIP–qPCR), and rescue experiments were conducted to verify the functional interactions among NPC1L1, Cyp27a1, 27‐OHC production, and the transcription factor C/EBPα. NPC1L1 expression was downregulated during osteogenesis, and its knockdown significantly enhanced osteogenic differentiation, proliferation, and migration. At the molecular level, NPC1L1 promoted cholesterol metabolism independently of its transport function, resulting in elevated 27‐OHC levels through increased expression of Cyp27a1. Elevated 27‐OHC suppressed osteogenesis through the induction of oxidative stress and the downregulation of osteogenic biomarkers (ALP, OPN, OSX, and OCN). In OVX mice, NPC1L1 knockdown significantly reversed osteoporosis‐related bone loss, as evidenced by improved trabecular parameters (BV/TV%, Tb.Th, Tb.N). Furthermore, we identified C/EBPα as a transcriptional activator of Cyp27a1, which mediates the regulatory effects of NPC1L1 on 27‐OHC production. NPC1L1 inhibits osteogenesis and contributes to OP by promoting the Cyp27a1‐dependent synthesis of 27‐OHC through the transcription factor C/EBPα. Targeted modulation of the NPC1L1‐C/EBPα‐Cyp27a1‐27‐OHC axis could provide novel therapeutic strategies for OP.https://doi.org/10.1096/fba.2025-0004427‐hydroxycholesterolcholesterol metabolismNPC1L1osteogenic differentiationosteoporosis |
| spellingShingle | Bohao Li Wuling Zhou Yueming Yu Boyu Chen Zhicheng Lv Jiarui Zhang Tieqi Zhang Shiwei Sun Lei Zhou Minghai Wang NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss FASEB BioAdvances 27‐hydroxycholesterol cholesterol metabolism NPC1L1 osteogenic differentiation osteoporosis |
| title | NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss |
| title_full | NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss |
| title_fullStr | NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss |
| title_full_unstemmed | NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss |
| title_short | NPC1L1 Drives Osteoporosis by Activating the C/EBPα/Cyp27a1/27‐Hydroxycholesterol Axis: A Novel Therapeutic Target for Bone Loss |
| title_sort | npc1l1 drives osteoporosis by activating the c ebpα cyp27a1 27 hydroxycholesterol axis a novel therapeutic target for bone loss |
| topic | 27‐hydroxycholesterol cholesterol metabolism NPC1L1 osteogenic differentiation osteoporosis |
| url | https://doi.org/10.1096/fba.2025-00044 |
| work_keys_str_mv | AT bohaoli npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT wulingzhou npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT yuemingyu npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT boyuchen npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT zhichenglv npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT jiaruizhang npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT tieqizhang npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT shiweisun npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT leizhou npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss AT minghaiwang npc1l1drivesosteoporosisbyactivatingthecebpacyp27a127hydroxycholesterolaxisanoveltherapeutictargetforboneloss |